Fig. 4

Knockdown of bmp4 negatively regulates shha expression and promotes SMC proliferation. bmp4 morphants (Bmp4 MO; A, B) and dorsomorphin treated embryos (C, D) show mild pericardial edema at 96 hpf (B, D) as compared to wild type embryos (A, C). At 96 hpf bmp4 morphants or dorsomorphin treated embryos show an underdeveloped gut (boxed area, B, D) as compared to wild type embryos (A, C). (E) Loss of Bmp4 leads to strong down-regulation of both shha and ihha at 96 hpf by qPCR (* refers to p<0.05; ** refers to p<0.01; n=3) as compared to wild type. Reduced shha and ihha expression is also seen in bmp4 morphants and dorsomorphin treated embryos as compared to wild type (F–M). (N–Q) The reduction in staining is also evident in longitudinal section. (R) The percentage of EDU positive smooth muscle and epithelial cells was quantified in wild type and bmp4 morphant embryos. There is no significant change of the percentage of EdU positive cell in the epithelium, but a significant upregulation of EDU positive smooth muscle cells (SMC). (S,T) Transverse sections of EdU stained cells (red) in the gut of 96 hpf embryos, counterstained with DAPI (blue). Proliferative SMCs are indicated by arrows. The magnification for A–D is shown in D, for F–M is in M, for N–Q is in Q, and for S, T is in T.